The UOK 257 cell line: a novel model for studies of the human Birt-Hogg-Dubé gene pathway

Abstract

The establishment, characterization, and tumorigenicity of a new epithelial cell line (UOK 257) derived from human renal carcinoma of an individual with Birt-Hogg-Dubé (BHD) syndrome are reported. Unlike other established renal tumor cell lines from sporadic renal cell carcinoma, this is the first established renal tumor cell line of BHD, an inheritable neoplastic syndrome. The isolated tumor cells display loss of contact inhibition in vitro, and produce subcutaneous tumors in mouse xenografts. Histopathologic, ultrastructural, and cytogenetic characterizations of the established tumor cells are reported. Cytogenetic analysis using spectral karyotyping on UOK 257 cells revealed 17p loss and a near-triploid and aneuploid karyotype with multiple fluorescence in situ hybridization analysis using a locus-specific gene probe for MYC. The result demonstrates that the established tumor cells consist of two cell populations, one containing four and one containing five copies of the MYC oncogene.

Fig. 1

(A–D). Light microscopic appearance with different histologies of kidney tumor mass from the patient with Birt-Hogg-Dubé ( BHD ) syndrome. (A) Large areas composed of nests of clear cells with enlarged, round to irregular nuclei with prominent nucleoli and abundant vacuolated cytoplasm, scattered atypical “naked” nuclei (x400) , (B) papillae lined by atypical epithelial cells with granular eosinophilic cytoplasm (x400) , ( C) atypical lining of the tubular structures (x400) , (D) histologically reminiscent foci of chromophobe renal cell carcinoma (x400) . (E–H) Tumor xenografts revealed similar histologic features compare with the case pathologic report. (E) Clear cell areas throughout the tumor (x400). (F) Papillary architecture (x200). (G) Areas with histologic resemblance to chromophobe (x200). (H) Solid areas of cells with eosinophilic cytoplasm, junction with complex of different histologies. (x200).

Fig. 1

(A–D). Light microscopic appearance with different histologies of kidney tumor mass from the patient with Birt-Hogg-Dubé ( BHD ) syndrome. (A) Large areas composed of nests of clear cells with enlarged, round to irregular nuclei with prominent nucleoli and abundant vacuolated cytoplasm, scattered atypical “naked” nuclei (x400) , (B) papillae lined by atypical epithelial cells with granular eosinophilic cytoplasm (x400) , ( C) atypical lining of the tubular structures (x400) , (D) histologically reminiscent foci of chromophobe renal cell carcinoma (x400) . (E–H) Tumor xenografts revealed similar histologic features compare with the case pathologic report. (E) Clear cell areas throughout the tumor (x400). (F) Papillary architecture (x200). (G) Areas with histologic resemblance to chromophobe (x200). (H) Solid areas of cells with eosinophilic cytoplasm, junction with complex of different histologies. (x200).

Fig. 2

Morphology of UOK 257 at 50% confluence (A) appearance small patched islands, at 90% (B) confluence, loss of contact inhibition is evident. From flow cytometry, the scatterplot (C) and its one-dimensional histogram projection depict the cell cycle distribution of the tumor cell population.

Fig. 3

(A–D) Ultrastructural features of findings from cultured UOK 257 tumor cells (Thin section (90nm) from fixed cells pellet) (X 6,500). (A): The arrows showed microvilli on one (apical) surface and basal lamina on the opposite surface. (B): Basal lamina –like material was focally abundant in between tumor cells. (C): Abundant mitochondria have appeared, consistent with oncocytic change. (D): Cytoplasmic microvesicles (arrowheads) were present in chromophobe renal cell carcinoma and rare tumor cells.

Fig. 4

A. A representative metaphase spread from cell line UOK 257 at passage18. (B). Composite karyotype of UOK 257 is near triploid, displaying multiple unbalanced translocations and deletions of chromosomes (white arrows).Presented to the left are inverted-DAPI stained chromosomes. The pseudo-colored chromosomes to the right were hybridized with SKY probes.

Fig. 5

(A) Two clones of UOK 257 are revealed by SKY and FISH, one with three copies of chromosome 8 and the other with four copies of chromosome 8 found in 50% of cells. (B) Fluorescence in situ hybridization using a locus-specific probe for the MYV oncogene (yellow signal) and whole chromosome paint for chromosome 8 (red). In addition, SKY analysis revealed that 8q21->8q24 (white arrows-MYC) is fused to the distal region of the unbalanced translocation, der(6)t(6;17)t(6;8).